Long term complications of diabetes can be reversed by intensive insulin therapy, however a major obstacle is the development of hypoglycemia. The adverse effects of hypoglycemia predominantly involve brain function, particularly cognition. Our goal is to better understand the consequences of CNS hypoglycemia and how it affects normal brain metabolism in diabetes and under intensive insulin treatment. The experiments described in this proposal are going to address the fundamental mechanisms underlying brain energy substrate utilization during hypoglycemia. Preliminray data our group has generated from type 1 diabetic patients suggests that during hypoglycemia uptake and metabolism of fuels other than glucose are increased. Based on this we want to assess the ability of short and medium chain fatty acids to support brain metabolism under hypoglycemia.We are interested in the effect of antecedent recurrent hypoglycemic episodes in diabetic and normal animals on rates of metabolism and blood brain barrier uptake of the alternate fuels lactate, acetate and octanoate. The respective contribution of these substrates to brain oxidative capacity in rodent models of diabetes and hypoglycemia have not yet been fully characterized. We are going to address this question in our first aim via MR spectroscopy in vivo, where we will look at the influence of recurrent hypoglycemia on metabolic flux rates of acetate and octanoid and associated glutamate/ glutamine cycling in the neuronal and astrocytic compartments. We will investigate in our second aim whether recurrent hypoglycemia in normal and diabetic rats induces an increase in blood brain barrier uptake of the alternate substrates lacate, acetate and octanoid. A potential underlying molecular mechanism to increased monocarboxylic acid transport could be the upregulation of transporter proteins. By measuring the mRNA and protein levels of monocarboxylic acid transporters 1, 2 and 4 by in situ hybridisation and western blot we will be able to answer this question. Whether alternate fuels, changes in substrate utilisation and increased metabolism are indeed able to sustain normal brain activity under hypoglycemia will be tested in aim three in a functional assay using EEC as well as by MR spectroscopy looking for high-energy phosphates. The long term goal of this proposal is to better understand the changes to transport and metabolism of alternate fuels that occur within the brain during hypoglycemia and how diabetes and intensive insulin treatment affect this process. This will provide the basis for the development of novel therapeutic approaches that could be used in clinical practice to protect the brain from hypoglycemia-induced injury.